The repair of large bone defects resulting from trauma, metabolic disorders, and tumor removal is a major medical challenge. Typically, such defects are treated with a bone allograft, where the terms “allograft” or “allogeneic transplant” are used interchangeably to refer to situations in which transplanted cells, tissues, or organs are sourced from a genetically non-identical member of the same species. However, allografts lack osteoinductive factors necessary to accelerate new bone growth and may carry the risk of disease transmission, since such grafts typically are harvested from cadavers. Due to these limitations, alternative strategies are needed.
Tissue engineering is one approach to the repair of large bone defects that has gained considerable interest. Tissue engineering is the application of principles and methods of engineering and life sciences toward a fundamental understanding and development of biological substitutes to restore, maintain and improve human tissue functions. Bone regeneration may be achieved by the use of osteogenic cells and/or factors to induce bone growth in combination with an appropriate scaffold to guide and support the laying down of new bone tissue. Optimally, a scaffold for bone tissue engineering should satisfy the following minimum requirements: biocompatibility (meaning the ability to coexist with living tissues or organisms without causing harm), osteoconductivity (meaning the ability to serve as a scaffold or matrix on which bone cells may attach, migrate and form new bone), porosity (meaning having minute openings, pores or holes that may be filled (permeated) by water, air or other materials), biodegradability (meaning having the ability to break down into harmless substances by the action of living organisms) and mechanical integrity (meaning having the ability to hold together and withstand chemical, physical, and biological forces over time).
The term “bioceramic” refers to ceramic materials employed within the body. Bioceramics employed within the body may be inert (meaning they remain unchanged), resorbable (meaning they dissolve) or active (meaning they may take part actively in physiological processes). Bioceramics may take many forms, including, but not limited to, microspheres, thin layers or coatings, porous networks, composites having a polymer component, and large well-polished surfaces. Direct use of ceramics for clinical applications has been limited because of their brittleness and difficulty in shaping.
Generally, those of skill in the art combine one ceramic and one polymer to create scaffolds appropriate for bone tissue engineering. K. Rezwan et al., Biodegradable and bioactive porous polymer/inorganic composite scaffolds for bone tissue engineering, Biomaterials 27 (2006) 3413-3431.
Several methods have been used to synthesize tissue engineering scaffolds based on polycaprolactone and polycaprolactone-tricalcium phosphate composites. For example, Zhou et al (In vitro bone engineering based on polycaprolactone and polycaprolactone-tricalcium phosphate composites, Polym Int 56 (2007) 333-342) used a fused deposition modeling method to synthesize a composite of poly(ε-caprolactone) (PCL) and tricalcium phosphate (TCP). Although a foaming method has been used to create hydroxyapatite-poly(L-lactic acid) ((HA)-(PLLA)) and β-TCP-poly(lactic acid) (β-TCP-PLA) composites M. Montjovent et al. Biocompatibility of bioresorbable poly(L-lactic acid) composite scaffolds obtained by supercritical gas foaming with human fetal bone cells, Tissue Engineering 11 (2005) 1640-1649, the authors did not create a composite of PLA, HA, and β-TCP together. G. Georgiou et al., Polylactic acid phosphate glass composite foams as scaffolds for bone tissue engineering, J. Biomed. Mat. Res. Part B: Applied Biomaterials, published online Jul. 12, 2006, used a foaming or compression molding method to synthesize a composite of PLA and a phosphate. See also, U.S. Pat. No. 5,626,861; U.S. Pat. No. 5,681,873, U.S. Pat. No. 5,766,618 (the '618 patent), U.S. Pat. No. 5,955,529; U.S. Pat. No. 6,165,486; U.S. Pat. No. 6,306,424; U.S. Pat. No. 6,730,252; U.S. Pat. No. 7,012,106; U.S. Pat. No. 7,022,522. None of these contemplate the use of electrospinning as a method to synthesize a scaffold containing ceramic polymer composites.
Electrospinning, another method that has been used to synthesize polymeric tissue engineering scaffolds, applies a high voltage to an ejectable polymer solution. The basic principle behind this process is that an electric voltage sufficient enough to overcome the surface tension of a polymeric solution causes the polymer droplets to elongate so that the polymer is splayed randomly as very fine fibers, which when collected on a grounded metal plate, form non-woven mats. Traditionally, electrospinning has yielded nonwoven mats (also called matrices and scaffolds) of nanometer sized fiber diameters and nanometer sized pore diameters. However, in order for cells to infiltrate into a scaffold and proliferate, micron sized fiber diameters and micron sized pore diameters are optimal. Since the diameter of a cell is approximately 10 μm to 20 μm, pore sizes at the cellular level or above are needed to allow for cell infiltration.
Polymer and calcium phosphate ceramic composites used in conventional scaffold-forming techniques are not easily adaptable to the electrospinning method. The parameters of voltage, flow rate, needle gauge size, distance to collection plate, and polymer solution concentration during processing need to be optimized to achieve fibrous mats. When combining a polymer with a ceramic in solution, in addition to optimizing these parameters, the homogeneity of the polymer-ceramic mixtures must be ensured. Moreover, the literature in this field does not provide sufficient guidance to enable one of skill in the art of tissue engineering to adapt polymer and ceramic composites to the electrospinning method using routine experimentation.
Previous work to develop scaffold materials for tissue engineering by electrospinning using polycaprolactone (PCL) or hydroxyapatite (HA) has produced mats containing nanometer sized fiber diameters having nanosized pore diameters in the mat. Such mats are not optimal for osteogenesis, because these pore diameters are below the preferred range of pore sizes for cell infiltration. See e.g. H. Yoshimoto et al., A biodegradable nanofiber scaffold by electrospinning and its potential for bone tissue engineering, Biomaterials 24 (2003) 2077-2082; M. Shin et al., In vivo bone tissue engineering using mesenchymal stem cells on a novel electrospun nanofibrous scaffold, Tissue Engineering 10 (2004) 33-41; C. Li et al., Electrospun silk-BMP-2 scaffolds for bone tissue engineering, Biomaterials 27 (2006) 3115-3124. HA also has been electrospun with PCL alone (P. Wutticharoenmongkol et al., Preparation and characterization of novel bone scaffolds based on electrospun polycaprolactone fibers filled with nanoparticles, Macromol. Biosci. 6 (2006) 70-77) and with PCL and collagen (J. Venugopal et al., Biocomposite nanofibres and osteoblasts for bone tissue engineering, Nanotechnology 18 (2007) 1-8).
Likewise, a number of patents have disclosed an electrospinning method for production of polymer nanofibers. These nanofiber mats have nanosized pore diameters in the mat, which are below the desired range of pore sizes necessary for cell infiltration. See, for example, U.S. Pat. No. 6,689,166; U.S. Pat. No. 6,790,528; U.S. Published Pat. App. No. 2004/0018226; U.S. Published Pat. App. No. 2006/0204539; U.S. Published Pat. App. No. 2006/0128012.
In order for a biodegradable scaffold to be successful, the material must have a rate of degradation that is commensurate with the growth of new bone. Ideally, the scaffold should degrade slowly enough to maintain structural support during the initial stages of bone formation, but fast enough to allow space for continuous growth of new bone. Previous studies have demonstrated the potential of biphasic compositions of HA and β-TCP ceramics for bone tissue engineering applications. One major advantage is that their rate of degradation correlates with bone tissue formation.
The present invention, which addresses this problem, provides compositions and methods of preparing a three-dimensional matrix of micron sized electrospun fibers, wherein the electrospun fibers are formed from a electrospun composite comprising a bioactive ceramic component and a polymer component. The matrix provides an osteoconductive and osteoinductive scaffold supporting osteogenesis and thereby may facilitate bone repair.